Avaliação da atividade imunomoduladora da fisalina F em células humanas e de camundongos

Abstract

Immune system disorders are responsible for the development of various diseases. The drugs used to treat these conditions have serious side effects, which creates the need for new pharmaceuticals. In this context, physalins have several pharmacological activities already described in the literature, with physalin F being an important compound with immunomodulatory activity. Therefore, the aim of this study was to investigate the immunomodulatory effects of physalin F in splenocytes from BALB/c mice and in human peripheral blood mononuclear cells (PBMCs). Initially, a cytotoxicity assay was performed on BALB/c mouse splenocytes and PBMCs. Subsequently, the effect of treatment on mitogen-activated lymphoproliferation was evaluated by CFSE or Celltiter Glo® labeling, and the effect of the compound on the production of inflammatory mediators was assessed by ELISA or ELISPOT. The interference of physalin F in the cell cycle and cell death patterns were evaluated by flow cytometry. The combined therapy of physalin F with dexamethasone was investigated using Celltiter Glo®. To explore the potential mechanisms of action, the effect of the compound on calcineurin activity was assessed using a calcineurin cellular activity assay kit. The in vivo effect was evaluated in a delayed-type hypersensitivity model. The molecule's interference in gene expression was evaluated by RT-PCR or PCR array. Additionally, molecular docking between physalin F and calcineurin was evaluated. Physalin F, at the tested concentrations (0.5, 1, and 2 µM), did not show cytotoxicity in BALB/c mouse splenocytes or human PBMCs. After treatment with physalin F, lymphoproliferation was inhibited, showing a synergistic effect with dexamethasone. Treatment with physalin F reduced the inflammatory mediators IL-2, IFN-γ, IL-10, IL-4, and IL-13 and decreased IFN-γ-producing cells. The treatment also induced a G1-phase cell cycle arrest and cell death by apoptosis. Physalin F reduced calcineurin activity and decreased paw edema in a delayed-type hypersensitivity model. The compound modulated key genes in the NFAT pathway, such as calcineurin, calmodulin, and NFAT1. The results demonstrate a potent immunosuppressive activity of physalin F.